Quick glimpse


We envision to keep fostering on continuous integration and development of highly reproducible workflows.

iMAP 1: Getting started workflow




iMAP 2: Microbiome Bioinformatics

Mothur Bioinformatics snakemake graph



16S classification options

1. Operational Taxonomic Units (OTUs)

  • OTUs are clusters of similar sequences and are commonly accepted as analytical units in microbial profiling when using 16S rRNA gene markers.

2. Phylotype

  • A phylotype in microbiome research is a DNA sequence or group of sequences sharing more than an arbitrarily chosen level of similarity of a 16S rRNA gene marker.

3. Amplicon Sequence Variant (ASV)

  • An ASV in microbiome research is any inferred single DNA sequences recovered from a bioinformatics analysis of 16S rRNA marker genes.
  • ASV is typically really a cluster of sequences that are one or two bases apart from each other.

4. Microbial Phylogenies

  • Microbial phylogenies are from gene sequence homologies. Models of mutation determine the most-likely evolutionary histories.



iMAP 3: Data tidying & transformation




iMAP 4 : Data analysis & visualization




iMAP 5 : Microbiome Machine Learning




Appendix

Reference Databases

  1. Mothur-based SILVA reference files4
  2. .
  3. Mothur-based RDP reference files5. Note: The RDP database is to classify 16S rRNA gene sequences to the genus level.
  4. ZymoBIOMICS Microbial Community Standard (Cat # D6306)6. The ZymoBIOMICS Microbial Community DNA Standard is designed to assess bias, errors and other artifacts after the step of nucleic acid purification.



Mermaid template

Troubleshooting (in progress)

  1. Are chimeras removed by default in newer versions?
    • Yes. Chimeras are removed by default. You can still run the remove.seqs command without error, but it is not necessary. Remove chimera sequence explained here
    .
  2. Mothur dist.seqs taking too long.
    • Merged reads are too long, probably over 300pb.
    • Reads not overlaping when merging the paired reads.
    • Too many uniques representative sequences probably caused by lack of overlapping.
    • No enough computer power which suggest a use of HPC or Cluster.



References

1. Köster, J., Mölder, F., Jablonski, K. P., Letcher, B., Hall, M. B., Tomkins-Tinch, C. H., Sochat, V., Forster, J., Lee, S., Twardziok, S. O., Kanitz, A., Wilm, A., Holtgrewe, M., Rahmann, S., & Nahnsen, S. (2021). Sustainable data analysis with snakemake. F1000Research, 10. https://doi.org/10.12688/f1000research.29032.2
2. Snakemake. (2023). Snakemake. https://snakemake.readthedocs.io/en/stable
3. Close, W. L. (2020). Mothur 16S v4 analysis pipeline. https://github.com/wclose/mothurPipeline
4. Mothur-based silva reference files. https://mothur.org/wiki/silva_reference_files/
5. Mothur-based RDP reference files. https://mothur.org/wiki/rdp_reference_files/
6. ZymoBIOMICS microbial community DNA standard (cat # D6306). https://www.zymoresearch.com/zymobiomics-community-standard